The present invention pertains to a novel medium adapted for the growth of Lactobacilli, which comprises a milk-derived base supplemented by at least four amino acids, ribonucleotide precursors, and iron. In particular, the present invention relates to the use of said novel medium for cultivating a variety of different Lactobacillus strains, e.g. L. johnsonii, L. acidophilus, L. gallinarum, etc. for the preparation of dairy products.
Lactic acid bacteria have been used since long for the production of a variety of food material, such as yoghurt, cheese, curd etc. In addition to their general usage in the food industry for fermentation purposes quite recently some strains belonging to the genus Lactobacillus or Bifidobacteria have attracted a great deal of attention due to probiotic properties attributed to them. Consequently, there was a desire to improve cultivation conditions so as to be able to maximize the growth and output of the microbial biomass.
One difficulty with large scale cultivation and application of lactic acid bacteria is their different nutritional requirements. In this context, different strains belonging to one specific genus or species require different media for optimal growth, which makes a standardized production of microbial biomasses complicated and cumbersome. For example, in producing a biomass of different strains of the genus Lactobacillus, a variety of different media have to be utilized, each of which does, however, only fulfil the nutritional needs of one particular strain, while not providing proper nutrients and/or environment for sufficient growth of other Lactobacillus strains therein.
A medium often utilized for cultivating lactic acid bacterial strains is cow milk. On the one hand, this medium provides a complex natural environment and its fermentation products, e.g. yoghurt, may be directly used as food material. Yet, this medium has proved to only support the growth of a limited number of strains of lactic acid bacteria. For example, Lactobacilli of the Johnson""s group A and B have been demonstrated to be essentially unable to proliferate and grow in milk, which makes this medium useless for said strains.
In some cases, bacterial growth could be improved when substances of an undefined and very complex composition, such as yeast extract or peptones of various origins, are added to the milk. However, these additional components can often cause an off-flavor with the result that cultures growing in a medium supplemented in such a way may not be used for industrial manufacture of dairy products. Moreover, the costs involved and the sometimes varying results in repeatability of achievable bacterial counts, makes such a process unsuitable for a commercial manufacture of these microbial strains.
In view of this, a problem of the present invention is to provide a medium which supports the growth of Lactobacillus strains while avoiding the shortcomings of the art.
The present invention solves the shortcomings of the art by providing a medium for the growth of Lactobacillus strains. The medium comprises a milk-derived base that has been supplemented by an additive system that includes at least four amino acids, at least two ribonucleosides or other ribonucleotide precursors, and beneficially iron. These components are added in amounts sufficient in combination to promote growth of lactobacilli in the medium. The specific amount of each of the ribonucleotide precursors, i.e., free bases, ribonucleosides, and deoxyribonucleosides, to be added to the base lie in the range from about 10 to about 500 milligrams per liter (mg/l) base, preferably from about 10 to about 100 mg/l base. Preferably, iron is added to the base in an amount ranging from about 10 to about 200 mg/l and more preferably in an amount from about 50 to about 100 mg/l milk.
Advantageously, the base is supplemented by at least four amino acids. The amino acids may be selected from any known or existing amino acid available to the skilled person. The amount of amino acids to be added to the milk base lies in the range of from about 10 to about 200 mg/l, preferably from about 50 to about 100 mg/l milk. In a preferred embodiment, the amino acids to be used are selected from the group consisting of cysteine, alanine, serine and isoleucine, which are found to particularly improve the growth conditions for Lactobacilli.
According to yet another preferred embodiment the base may also be supplemented by compounds providing antioxidant or reducing activity. Examples of such compounds include ascorbic acid, vitamin E, tocotrienol, ubiquinol, 13-carotene and other carotinoids, rosemary compounds (e.g. camosol) and other flavonoids, and other sulphur containing antioxidants including glutathione, lipoic acids, N-acetylcysteine or compounds bearing suithydryl groups, cysteine or thioglycollic acid, or mixtures thereof. Cysteine is preferred as an amino acid and as a compound providing reducing activity.